The present invention relates in general to cell culturing systems and specifically to a hollow fiber culturing system and/or ex vivo extracorporeal treatment means which circulates two different types of media, a gas and a liquid.
The most widely used cell culturing systems comprise batch culturing and hollow fiber culturing. Batch culturing comprises growing cells on the interior surface of plastic or glass rollers, or attaching the cells to a flat surface on stationary containers such as petri dishes.
In contrast, hollow fiber culturing systems allow for the in vitro growth of cells on semi-permeable tubular membranes or capillaries. The cells attach to the outer surface of the capillary walls. A nutrient media is circulated through the capillaries to diffuse from the perfusing medium though the capillary walls to the cells. In return, cell products diffuse from the cells through the capillary walls and into the perfusate from which they can be harvested if one so desires. While the prior art is replete with hollow fiber cell culturing systems an in vitro culturing system which is identical to an in vivo system is still illusive Generally speaking, hollow fiber culturing systems encounter three major categories of problems. First, the cells must be supplied with a nutrient media. Second, an optimal and constant environment must be maintained while the cells are metabolizing. Third, a suitable substrate must be available for cell attachment.
One, of the most significant problems within the nutrient category to be overcome in the production of tissue-like derivatives for creating artificial organs via hollow fibers, is oxygenation. Without an adequate oxygen supply, cells cannot metabolize i.e. regulate themselves and grow. The prior art culture systems have supplied oxygen with preliminary perfusion. This oxygenating of the fluid, from outside the fibers is inadequate in that cells prefer a continuous and steady rate of oxygen. Moreover, it is best to supply it directly to the cells and not a distance away. This again is only a mimicking of in vivo conditions where the blood with its hemoglobin carrying molecules bathes cells. In sum, the prior art hollow fiber systems fall short of xe2x80x9cbathingxe2x80x9d the cells by either supplying an initial non-continuous oxygen perfusion or by doing so at a distance away from the cells.
The prior art has also tried perfusing the media directly with oxygen. However, aqueous nutrient media equilibrated with air is only able to carry 4.5 ml. of oxygen per liter (37xc2x0 C. 760 mm of Hg). The inability of aqueous solutions to carry oxygen results in oxygen perfusion being the rate limiting step in the culturing process. In order to overcome oxygen shortages the rate must be increased. High circulation rates result in high internal pressures and turbulence. Since cells are quite fragile, vigorous aeration prevents cell propagation and may lead to the denaturation of proteins.
In conclusion, neither pre-oxygenation and/or oxygenating the exterior of the capillaries and/equilibrating the media with air is not optimal. The problem is further compounded when artificial organs are the goal, since all the cells are in a catabolic state and require an even greater amount of oxygen. A discontinuous and/or inadequate and/or turbulent oxygen supply results in a range of cellular problems ranging from irregular cell metabolism and growth to premature death. Thus it is a primary objective of this invention to provide for optimal oxygenation.
Given the aforementioned, it is best to separate the oxygen supply from the medium support. The prior art devices which have done so, employ both a hollow fiber diffusion mechanism and a bath. Either oxygen is diffused through the hollow fiber and the cells are bathed in the medium or the cells are perfused with oxygen and the medium is diffused through the hollow fiber. While this system is quite advantageous, it is difficult to remove products when the hollow fiber is only used for oxygen supply. Conversely, it is problematic for supplying a continuous and steady supply of oxygen, when the hollow fiber only perfuses media and not oxygen. Thus it is a further objective of the present invention to provide for a device which perfuses both a gas and a liquid.
U.S. Pat. No. 4,184,922 employs two perfusion circuits. However, both circuits are perfused with liquid media. It does not perfuse a gas. The patent therefore does not address oxygenation nor improve oxygenation beyond that of the prior art. Moreover, given the dual liquid perfusion, the patent only employs ultrafiltration fibers. The present invention may still be further differentiated in that the dimensions of the present invention""s unit range from 7 to 40 cm in length and 1.4 to 5.0 cm in diameter versus the 5 cm long and 0.26 cm diameter unit described in U.S. Pat. No. 4,184,922. Lastly the system described in the aforementioned patent is limited to 50 fibers whereas the present invention may employ up to 7500 fibers thereby increasing the desired product by an extremely advantageous margin. Further advantages of the present invention will become apparent in the remaining specification.
In sum, it is highly desirable to produce a hollow fiber cell culturing system and/or ex vivo extracorporeal blood, plasma and/or serum, treatment unit which overcomes the aforementioned prior art disadvantages.
The task of the present invention can be solved in an elegant and novel manner by providing for a hollow fiber system wherein some fibers carry a gas and other fibers carry a liquid. More specifically, by providing for a cell culture unit comprising a shell having two ends with a liquid and a gas perfusion port located on each of said end, said ends defining an elongated chamber therebetween. Said cell culture unit further comprising liquid perfusion fibers connected to said liquid perfusion ports and gas perfusion fibers connected to said gas perfusion ports; said fibers defining extracapillary and intracapillary space. Still further said unit comprising means communicating with the intracapillary space of said gas perfusion fibers, means communicating with the intracapillary space of said liquid perfusion fibers and means communicating with said extracapillary space.